This invention relates to cancer therapy. More particularly, this invention relates to the modulation of multiple drug resistance in cells by antisense oligonucleotides complementary to portions of the MDR-1 gene.
Chemotherapy has been successfully used to control metastatic cancers which cannot be cured by surgery because of their widespread dissemination throughout the body. However, the effective use of chemotherapy is limited by inherent and acquired resistance of many tumors to a variety of structurally unrelated chemotherapeutic drugs.
One mechanism of drug resistance involves the overexpression of a 170,000 dalton membrane protein, P-glycoprotein (or P170), which acts as an ATP-dependent, chloride-selective efflux pump for a variety of hydrophobic chemotherapeutic drugs (Roninson (1991) Molecular and Cellular Biology of Multi-Drug Resistance in Tumor Cells, Plenum Press, NY; Valverde et al. (1992) Nature 355: 830-833). This protein is encoded by the multidrug resistance-1 gene (MDR-1) gene. A recent estimate suggests that expression of MDR-1 will result in greater than 250,000 new cancers displaying clinically significant resistance (Gottesman (1993) Cancer Res. 53: 747-754).
Reversal or suppression of the resistant phenotype has been accomplished in cell culture systems and in animal experiments with small molecule inhibitors of P170, thereby rendering chemotherapeutic drugs more effective at killing tumor cells (Tsuruo et al. (1981) Cancer Res. 41: 1967-1972). For example, verapamil, cyclosporin, and trifluoroperazine have been used to reverse the MDR phenotype (see, e.g., Thierry et al. (1993) Biochem. Biophys. Res. Commun. 190: 952-960; and Jaroszewski et al. (1990) Cancer Commun. 2: 287-294). However, the clinical use of many of these drugs is limited due to toxic side effects and lack of specificity (see, e.g., Ozols et al. (1987) J. Clin. Onc. 5: 641-647).
Immunoglobulins have also been used to modulate P170-mediated drug resistance. For example, it has been demonstrated that monoclonal antibodies and ricin-containing immunotoxins eradicate kidney carcinoma cell cultures (Efferth et al. (1993) Oncol. 50: 303-308). However, normal kidney cells expressing high levels of P-170 were similarly affected, indicating that clinical application of these antibodies and immunotoxins may be limited by toxic side effects on normal, P170-expressing tissues in cancer patients.
Recently, new chemotherapeutic agents have been developed which are capable of modulating cellular and foreign gene expression (see, Zamecnik et al. (1978) Proc. Natl. Acad. Sci. (USA) 75: 280-284). These agents, called antisense oligonucleotides, bind to target single-stranded nucleic acid molecules according to the Watson-Crick rule or to double stranded nucleic acids by the Hoogsteen rule of base pairing, and in doing so, disrupt the function of the target by one of several mechanisms: by preventing the binding of factors required for normal transcription, splicing, or translation; by triggering the enzymatic destruction of mRNA by RNase H, or by destroying the target via reactive groups attached directly to the antisense oligonucleotide.
Antisense oligonucleotides have been designed to inhibit the expression of MDR-1. For example, oligonucleotides complementary to the initiation codon of MDR-1 have been demonstrated to inhibit P170 expression (Rivoltini et al. (1990) Int. J. Cancer 46: 727-732; Vasanthakumar et al. (1989) Cancer Commun. 1: 225-232). Oligonucleotides complementary to the 5' end region of MDR-1 have been shown to decrease P170 synthesis and resistance to doxorubicin in various resistant cells (Corrias et al. (1992) Anticancer Res. 12: 1431-1438; Thierry et al. (1993) Biochem. Biophys. Res. Commun. 190: 952-960), and to inhibit the expression of P170 in kidney carcinoma and normal kidney cells (Efferth et al. (1993) Oncol. 50: 303-308). In addition, phosphorothioate oligonucleotides complementary to the 5' end of an intron/exon border, or to an EcoRI site have been demonstrated to inhibit P170 expression, but not necessarily with a decrease in the resistance phenotype.
There is, therefore, a need for better antisense oligonucleotides directed against MDR-1 that provide better diminution of the resistance phenotype.